The overall goal of this proposal is to obtain a better understanding of the nature and events occurring upon development of activated Natural Killer(NK) Cells and how these cells are involved in in vivo situations such as pregnancy, allograft rejection and immunotherapy. Specifically, the phenotypic and functional characterization of purified Mixed Lymphocyte Culture(MLC) induced NK cells and their precursor cells will be examined and compared to endogenous and interferon induced NK. The phenotypic characterization of purified MLC induced NK and their precursors will be performed using a large panel of well characterized monoclonal antibodies. These results will be correlated with those for endogenous and interferon activated NK previously published by us. In addition, the kinetics of expression and the role of MLC induced regulatory cells upon the expression of these antigens will be examiend. The functional characterization will be performed on the MLC induced NK as well as interferon activated NK and endogenous NK by using the single cell assay in conjunction with inhibitory monoclonal antibodies. These antibodies will be used as probes to examine tumor cell recognition and effector cell structures involved in the lytic process. Specifically, the examination of the site of action of these antibodies on uropod formation and function, membrane integrity and ultrastructure will be performed by light, EM, and flourescent microscopy and will provide much insight into the cell biology and mechanism of tumor cell lysis and recognition by endogenous and activated Nk cells. Further functional characterization will include the specificity of these cells on the single cell level and the investigation of various regulatory cells involved in the induction and regulation of MLC generated NK. This will be accomplished using FACS cell sorting of various regulatory populations. Finally, as an in vivo correlate to the in vitro MLC system different populations of lymphocytes from pregnant patients and cord blood will be examined for their regulatory role in the induction and expression of MLC and MLC-induced NK. In summary, these studies in addition to addressing basic questions such as heterogeneity, specificity, and mechanisms and regulation of tumor cell lytic processes by endogenous and activated NK, will also impact on immunotherapy, bone marrow and renal allograft rejection and monitoring, as well as immunosuppression seen in pregancy.